Integrated analysis of gut metabolome, microbiome, and exfoliome data in an equine model of intestinal injury.

BACKGROUND: The equine gastrointestinal (GI) microbiome has been described in the context of various diseases. The observed changes, however, have not been linked to host function and therefore it remains unclear how specific changes in the microbiome alter cellular and molecular pathways within the GI tract. Further, non-invasive techniques to examine the host gene expression profile of the GI mucosa have been described in horses but not evaluated in response to interventions. Therefore, the objectives of our study were to (1) profile gene expression and metabolomic changes in an equine model of non-steroidal anti-inflammatory drug (NSAID)-induced intestinal inflammation and (2) apply computational data integration methods to examine host-microbiota interactions. METHODS: Twenty horses were randomly assigned to 1 of 2 groups (n = 10): control (placebo paste) or NSAID (phenylbutazone 4.4 mg/kg orally once daily for 9 days). Fecal samples were collected on days 0 and 10 and analyzed with respect to microbiota (16S rDNA gene sequencing), metabolomic (untargeted metabolites), and host exfoliated cell transcriptomic (exfoliome) changes. Data were analyzed and integrated using a variety of computational techniques, and underlying regulatory mechanisms were inferred from features that were commonly identified by all computational approaches. RESULTS: Phenylbutazone induced alterations in the microbiota, metabolome, and host transcriptome. Data integration identified correlation of specific bacterial genera with expression of several genes and metabolites that were linked to oxidative stress. Concomitant microbiota and metabolite changes resulted in the initiation of endoplasmic reticulum stress and unfolded protein response within the intestinal mucosa. CONCLUSIONS: Results of integrative analysis identified an important role for oxidative stress, and subsequent cell signaling responses, in a large animal model of GI inflammation. The computational approaches for combining non-invasive platforms for unbiased assessment of host GI responses (e.g., exfoliomics) with metabolomic and microbiota changes have broad application for the field of gastroenterology. Video Abstract.

Bedside physician led US-guided supra-clavicular lymph node biopsy and ROSE (rapid on-site evaluation): SVC obstruction swift management in lung cancer.

Superior vena cava obstruction (SVCO) is an oncological emergency and can often be linked to an underlying lung malignancy. Due to the potential life-threatening risks associated with SVCO, it necessitates urgent diagnosis and management. In this report, we discuss 3 case studies where the use of ultrasound-guided supraclavicular lymph node biopsy was used to obtain a biopsy from patients with SVCO, followed by rapid on-site evaluation (ROSE). The benefits of this technique ensure a more rapid histological diagnosis, while also involving a less invasive procedure for the patient. The histological diagnosis is essential in improving patient outcomes when treating those with SVCO as the recommended treatments vary depending on the underlying type of lung malignancy. Having this information can help the clinician swiftly employ the optimal treatment pathway for the patient.

Rapid improvement in symptoms and physical function following ibrutinib initiation in chronic lymphocytic leukemia and the associated changes in plasma cytokines.

A prospective pilot study was carried out on 34 CLL patients treated with ibrutinib, evaluating the effects on symptoms and physical function with changes in plasma exosomes (EXs), ß2-microglobulin (ß2M) and 26 plasma cytokines. The revised Edmonton Symptom Assessment Scale (ESAS-R) demonstrated moderate fatigue, shortness of breath and a sense of unwellness before treatment, which significantly improved within 2 weeks of starting ibrutinib. These changes were associated with a rapid improvement in sit-to-stand and 4 m walking speeds. The plasma levels of CCL11, IL-7, -8 and -10 dropped initially while the levels of TNF-α/-ß, CCL3, CCL4, CCL17, and IL-16 continued to decline for 12 months. Despite the initial lymphocytosis, plasma ß2M levels fell but no consistent change in plasma EXs occurred. Thus, ibrutinib can produce a rapid and sustained improvement in symptoms and physical function in CLL, associated with a decline in multiple plasma cytokines.

I22: SAXS/WAXS beamline at Diamond Light Source - an overview of 10 years operation.

Beamline I22 at Diamond Light Source is dedicated to the study of soft-matter systems from both biological and materials science. The beamline can operate in the range 3.7 keV to 22 keV for transmission SAXS and 14 keV to 20 keV for microfocus SAXS with beam sizes of 240 µm × 60 µm [full width half-maximum (FWHM) horizontal (H) × vertical (V)] at the sample for the main beamline, and approximately 10 µm × 10 µm for the dedicated microfocusing platform. There is a versatile sample platform for accommodating a range of facilities and user-developed sample environments. The high brilliance of the insertion device source on I22 allows structural investigation of materials under extreme environments (for example, fluid flow at high pressures and temperatures). I22 provides reliable access to millisecond data acquisition timescales, essential to understanding kinetic processes such as protein folding or structural evolution in polymers and colloids.

Association of anthropometric status and residential locality factors with cognitive scores of 4-6-year-old children in Kwazulu-Natal, South Africa.

Background: The Asenze study has the long-term goal of promoting better physical, cognitive and psychosocial functioning of children in a rural area in KwaZulu-Natal, 50 km from Durban with a view to planning interventions to promote growth and development for very young children. The specific objective in this paper was to provide information for the Child Health and Development project of the Valley Trust to assist with intervention planning. The broader goal was to assess developmental delays in communities ravaged by the HIV epidemic.The Asenze study was designed in two phases from 2008 and 2012. The current paper reports on 1581 4-6years old children in the baseline phase (2008-2010) in the five adjacent tribal areas in the study area. Method: The participants included all the 4 - 6 year olds whose parents had consented to inclusion in the project and their caregivers. Data were derived from a brief questionnaire administered in the homes of participants, and subsequently from medical and psychological assessments of the children and their caregivers at the Asenze clinic. The association between child factors and other factors (geographic area, socioeconomic status (SES), parental level of education, the child's pre-school education) on the one hand and, the child's cognitive performance (as measured by the Grover Counter and subtests of the KABC-11) were analysed. Linear regression models were employed to determine which predictor variables of interest in a model were associated with the children's cognitive scores as the dependent variables. Results: Based on the data, the principal factors associated with children's cognitive outcomes were height-for-age z-score (HAZ), preschool education and the area of residence, Generally children who had low cognitive scores were more often stunted (as defined by the WHO anthropometric tables), had not had pre-school education, and came from areas less favourable in terms of local infrastructure and access to employment opportunities and arable land. Conclusion: The finding from this cross-sectional analysis of baseline data showed that in addition to height for age and pre-school education, which are commonly thought to impact on cognition, the local authority area where the children lived was associated with their scores on cognitive tests. This has implications for intervention planning. The functioning of local government in promoting the type of community development which will protect the rights of children should be taken into account.

Natural drugs, not so natural effects: Neonatal abstinence syndrome secondary to 'kratom'.

BACKGROUND: Mitragyna speciosa, also known as kratom, is obtained from the coffee plant family 'Rubiaceae.' Kratom is available in the form of capsules, whole, processed and powdered leaves, and as liquids. Secondary to its 'natural herb' status and opioid effects, it is misconceived to be a safe alternative for the treatment of chronic pain. The use of kratom has increased by tenfold in the United States since 2010. METHODS AND RESULTS: We report a term neonate who was born to a chronic kratom user and required treatment with opiates for neonatal drug withdrawal. CONCLUSION: Physicians should be aware of these herbal supplements and its potential withdrawal effects in newborn which cannot be picked up by the standard toxicology screen.

Selective method for identification and quantification of Bifidobacterium animalis subspecies lactis BB-12 (BB-12) from the gastrointestinal tract of healthy volunteers ingesting a combination probiotic of BB-12 and Lactobacillus rhamnosus GG.

AIM: To develop a novel validated method for the isolation of Bifidobacterium animalis ssp. lactis BB-12 (BB-12) from faecal specimens and apply it to studies of BB-12 and Lactobacillus rhamnosus GG (LGG) recovered from the healthy human gastrointestinal (GI) tract. METHODS AND RESULTS: A novel method for isolating and enumerating BB-12 was developed based on its morphologic features of growth on tetracycline-containing agar. The method identified BB-12 correctly from spiked stool close to 100% of the time as validated by PCR confirmation of identity, and resulted in 97-104% recovery of BB-12. The method was then applied in a study of the recovery of BB-12 and LGG from the GI tract of healthy humans consuming ProNutrients® Probiotic powder sachet containing BB-12 and LGG. Viable BB-12 and LGG were recovered from stool after 21 days of probiotic ingestion compared to baseline. In contrast, no organisms were recovered 21 days after baseline in the nonsupplemented control group. CONCLUSIONS: We demonstrated recovery of viable BB-12, using a validated novel method specific for the isolation of BB-12, and LGG from the GI tract of healthy humans who consumed the probiotic supplement. SIGNIFICANCE AND IMPACT OF THE STUDY: This method will enable more detailed and specific studies of BB-12 in probiotic supplements, including when in combination with LGG.

Evaluation of the Walking Index for Spinal Cord Injury II (WISCI-II) in children with Spinal Cord Injury (SCI).

STUDY DESIGN: Mixed methods were used in this study. The appropriateness of the levels of the Walking Index for Spinal Cord Injury II (WISCI-II) for application in children was critically reviewed by physical therapists using the Modified Delphi Technique, and the inter- and intra-rater reliability of the WISCI-II in children was evaluated. OBJECTIVES: To examine the construct validity, and to establish reliability of the WISCI-II related to its use in children with spinal cord injury (SCI). SETTING: United States of America. METHODS: Using a Modified Delphi Technique, physical therapists critically reviewed the WISCI-II levels for pediatric utilization. Concurrently, ambulatory children under age 18 years with SCI were evaluated using the WISCI-II on two occasions by the same therapist to establish intra-rater reliability. One trial was photographed and de-identified. Each photograph was reviewed by four different physical therapists who gave WISCI-II scores to establish inter-rater reliability. Summary and descriptive statistics were used to calculate the frequency of yes/no responses for each WISCI-II level question and to determine the percent agreement for each question. Inter- and intra-rater reliability was calculated using interclass correlation coefficients (ICCs) with 95% confidence intervals (CI). RESULTS: Construct validity was confirmed after one Delphi round during which at least 80% agreement was established by 51 physical therapists on the appropriateness of the WISCI-II levels for children. Fifty-two children with SCI aged 2-17 years completed repeated WISCI-II assessments and 40 de-identified photographs were scored by four physical therapists. Intra- and inter-rater reliability was high (ICC=0.997, CI=0.995-0.998 and ICC=0.97, CI=0.95-0.98, respectively). CONCLUSION: This study demonstrates support for the use of the WISCI-II in ambulatory children with SCI. SPONSORSHIP: This study was funded by the Craig H Neilsen Foundation, Spinal Cord Injury Research on the Translation Spectrum, Senior Research Award #282592 (Mulcahey, PI).

The experiences of survivors and trauma counselling service providers in northern Uganda: Implications for mental health policy and legislation.

Previous research in northern Uganda found high levels of trauma-related difficulties amongst the conflict-affected population. There is international evidence that psychological therapy can reduce depression, as one of the psychological effects of trauma, but very limited literature regarding the experiences of trauma counselling in Sub-Saharan Africa. The current British Academy and Leverhulme-funded research investigated the experiences of service users and providers of trauma services in Kitgum and Gulu, northern Uganda. It also examined their implications for mental health policy and legislation. A decision was made to utilise qualitative methodology to highlight the in-depth experiences of participants. The researcher's carried out interviews with 10 women and 10 men survivors attending trauma services in Kitgum and Gulu. The researchers also interviewed 15 key informants in Kitgum, Gulu and Kampala including trauma counselling service providers, ministers, cultural leaders and mental health professionals. The authors report the findings of the research based on thematic analysis of the interviews. Themes included the experiences of survivors, bearing witness and instilling hope, constraints to service provision, stigma and abuse, holistic approach, service providers doing their best, specialist populations, limited understanding, training and skills development, gaps in service provision and mental health policy and legislation. The interviews resulted in a clear indication that counselling and medication was valued by service users, and that service providers felt the treatments that were provided improved depression, and increased empowerment and engagement in social activities. However, the authors argue that there was a limit to the benefits that could be achieved without using the holistic approach that the survivors requested. Thus, in cases of trauma arising from conflict, there is a clear need for the state to ensure reparation and/or justice for the atrocities witnessed by and perpetrated against survivors. This might include the provision of compensation, which would help to meet social needs and reduce feelings of shame and anger.

Maternal and neonatal outcomes for pregnancies before and after gastric bypass surgery.

BACKGROUND: Interaction between maternal obesity, intrauterine environment and adverse clinical outcomes of newborns has been described. METHODS: Using statewide birth certificate data, this retrospective, matched-control cohort study compared paired birth weights and complications of infants born to women before and after Roux-en-Y gastric bypass surgery (RYGB) and to matched obese non-operated women in several different groups. Women who had given birth to a child before and after RYGB (group 1; n=295 matches) and women with pregnancies after RYGB (group 2; n=764 matches) were matched to non-operated women based on age, body mass index (BMI) prior to both pregnancy and RYGB, mother's race, year of mother/s birth, date of infant births and birth order. In addition, birth weights of 13 143 live births before and/or after RYGB of their mothers (n=5819) were compared (group 3). RESULTS: Odds ratios (ORs) for having a large-for-gestational-age (LGA) neonate were significantly less after RYGB than for non-surgical mothers: ORs for groups 1 and 2 were 0.19 (0.08-0.38) and 0.33 (0.21-0.51), respectively. In contrast, ORs in all three groups for risk of having a small for gestational age (SGA) neonate were greater for RYGB mothers compared to non-surgical mothers (ORs were 2.16 (1.00-5.04); 2.16 (1.43-3.32); and 2.25 (1.89-2.69), respectively). Neonatal complications were not different for group 1 RYGB and non-surgical women for the first pregnancy following RYGB. Pregnancy-induced hypertension and gestational diabetes were significantly lower for the first pregnancy of mothers following RYGB compared to matched pregnancies of non-surgical mothers. CONCLUSION: Women who had undergone RYGB not only had lower risk for having an LGA neonate compared to BMI-matched mothers, but also had significantly higher risk for delivering an SGA neonate following RYGB. RYGB women were less likely than non-operated women to have pregnancy-related hypertension and diabetes.

What will it take for recovery to flourish in Hong Kong?

The notion of mental health "recovery" is beginning to stimulate important changes in the mental health care provided to persons with serious mental illnesses in Hong Kong. However, the Chinese culture poses many challenges to implementing the types of recovery-oriented practices developed over the last 2 decades in the West. This article considers some of the challenges that policy makers, system leaders, practitioners, family members, and persons with mental illnesses themselves may face in attempting to transform care in Hong Kong. In addition to shifting from an individualistic to a more collectivist culture that emphasises the importance of family involvement, the primarily linear notion of mental stability that currently guides practice may need to be reconsidered in the face of evidence which suggests that recovery is a non-linear path that involves hard work both on the part of the individual as well as the family.

Variant CJD and blood transfusion: are there additional cases?

BACKGROUND AND OBJECTIVES: In this study, we compare variant Creutzfeldt-Jakob disease (vCJD) cases definitely linked to blood transfusion, those with a history of blood transfusion in which no donor has developed vCJD and primary cases with no history of blood transfusion. The aim is to determine whether there are any differences in the demographics or clinical phenotype in these groups that might suggest additional cases of transfusion transmission of vCJD. MATERIALS AND METHODS: All cases of vCJD who are old enough to donate blood (i.e. >17 years old) are notified to the UKBTS at diagnosis, regardless of whether they are known to have a blood donation history. A search is then made for donor records and, if found, all components produced and issued to hospitals are identified and their fate determined. Recipient details are then checked against the NCJDRSU register to establish whether there is a match between these individuals and patients who have been diagnosed with vCJD. In the reverse study, attempts are made to trace the donors to all cases reported to have received a blood transfusion and donors' details are checked against the register to determine if any have developed vCJD. RESULTS: Of the 177 cases of vCJD diagnosed in the UK as of 1 February 2014, the TMER study identified 15 cases reported to have received a blood transfusion. Transfusion records were unavailable for 4 of these cases, all pre-1980, and in one other case there was no transfusion recorded in the medical notes. Transfusion records were found for 10 cases. One case transfused at symptom onset was excluded from this analysis. The mean age at onset of symptoms of the remaining nine transfusion recipients (four female and five male) was 42·9 years; 57·6 years in the three known transfusion-transmitted cases and 35·5 years in the six not linked cases. In one of these cases, details of components transfused were unavailable, and the remaining five cases received a total of 116 donor exposures with 112 donors identified, none of whom is known to have developed clinical vCJD. To date, five of the 112 identified donors have died and none was certified as dying of vCJD or any other neurological disorder. Two of the transfusion-transmitted cases did not fulfil diagnostic criteria for probable vCJD during life but were confirmed at post-mortem. Both cases were in the older age range (68 and 74 years, respectively), and neither had a positive MRI brain scan. The remaining cases all fulfilled the criteria for the diagnosis of vCJD in life, but two of these had atypical features and were older than the expected age at onset for vCJD. CONCLUSION: In conclusion, it is possible that one or more of the vCJD cases that received a blood transfusion derived from an individual not known to have vCJD were infected by the blood transfusion. However, the evidence for this is weak, and the absence of a past history of transfusion in most cases of vCJD excludes a large number of unrecognised transfusion-transmitted cases.

Disruption of epithelial architecture caused by loss of PTEN or by oncogenic mutant p110α/PIK3CA but not by HER2 or mutant AKT1.

Genetic changes in HER2, PTEN, PIK3CA and AKT1 are all common in breast cancer and lead to the elevated phosphorylation of downstream targets of the PI3K/AKT signalling pathway. Changes in HER2, PTEN, PIK3CA and AKT have all been reported to lead to both enhanced proliferation and failures in hollow lumen formation in three dimensional epithelial culture models, but it is not clear whether these failures in lumen formation are caused by any failure in the spatial coordination of lumen formation (hollowing) or purely a failure in the apoptosis and clearance of luminal cells (cavitation). Here, we use normal murine mammary gland (NMuMG) epithelial cells, which form a hollow lumen without significant apoptosis, to compare the transformation by these four genetic changes. We find that either mutant PIK3CA expression or PTEN loss, but not mutant AKT1 E17K, cause disrupted epithelial architecture, whereas HER2 overexpression drives strong proliferation without affecting lumen formation in these cells. We also show that PTEN requires both lipid and protein phosphatase activity, its extreme C-terminal PDZ binding sequence and probably Myosin 5A to control lumen formation through a mechanism that does not correlate with its ability to control AKT, but which is selectively lost through mutation in some tumours. These findings correlate AKT-independent signalling activated by mutant PIK3CA or PTEN loss, but not strongly by HER2, with disrupted epithelial architecture and tumour formation.

Predictive value of bronchoscopy after infant cardiac surgery: a prospective study.

BACKGROUND AND AIMS: Airway evaluation following infant cardiac surgery often reveals evidence of tracheobronchial narrowing. We studied the association between airway narrowing and extubation failure (EF) in this population. METHODS: Prospective cohort study of infants (age ≤6 months) from March-September 2009. Flexible bronchoscopy (FB) evaluations were obtained using a standardised protocol after operative intervention. The primary endpoint was the development of extubation failure (EF; defined as the need for invasive mechanical ventilation ≤48 h after primary extubation) and several secondary endpoints. RESULTS: Fifty-three patients were evaluated at a median age of 81 [interquartile range (IQR) 13-164] days and weight of 4.2 (IQR 3.2-6.0) kg; 13 (25 %) of the patients had single ventricle palliations and two subsequently underwent heart transplantation. Significant airway narrowing was noted in 15 of 30 [50 %, 95 % confidence interval (CI) 31-69 %] patients who underwent FB; ten of the 53 patients (19 %, 95 %CI 10-32 %) subsequently developed EF. Narrowed airway calibre on bronchoscopy had a sensitivity and specificity of 50 % (95 %CI 28-71 %) and 50 % (95 %CI 28-71 %), respectively, for EF. The single greatest predictor of EF by univariate analysis was the need for preoperative ventilation [odds ratio (OR) 6.5, 95 %CI 1.3-33.2, p = 0.03]. Patients with EF had a greater likelihood of intensive care readmission (OR 4.8, 95 %CI 1.1-21, p < 0.04) during the same hospital admission. CONCLUSIONS: Airway narrowing on FB is noted frequently after infant cardiac surgery. Overall assessment and presence of narrowing on bronchoscopy had poor sensitivity and specificity for EF in our cohort. Expert assessment of tracheobronchial narrowing on FB has poor to moderate inter-rater reliability.

Serial analysis of the gut and respiratory microbiome in cystic fibrosis in infancy: interaction between intestinal and respiratory tracts and impact of nutritional exposures.

UNLABELLED: Pulmonary damage caused by chronic colonization of the cystic fibrosis (CF) lung by microbial communities is the proximal cause of respiratory failure. While there has been an effort to document the microbiome of the CF lung in pediatric and adult patients, little is known regarding the developing microflora in infants. We examined the respiratory and intestinal microbiota development in infants with CF from birth to 21 months. Distinct genera dominated in the gut compared to those in the respiratory tract, yet some bacteria overlapped, demonstrating a core microbiota dominated by Veillonella and Streptococcus. Bacterial diversity increased significantly over time, with evidence of more rapidly acquired diversity in the respiratory tract. There was a high degree of concordance between the bacteria that were increasing or decreasing over time in both compartments; in particular, a significant proportion (14/16 genera) increasing in the gut were also increasing in the respiratory tract. For 7 genera, gut colonization presages their appearance in the respiratory tract. Clustering analysis of respiratory samples indicated profiles of bacteria associated with breast-feeding, and for gut samples, introduction of solid foods even after adjustment for the time at which the sample was collected. Furthermore, changes in diet also result in altered respiratory microflora, suggesting a link between nutrition and development of microbial communities in the respiratory tract. Our findings suggest that nutritional factors and gut colonization patterns are determinants of the microbial development of respiratory tract microbiota in infants with CF and present opportunities for early intervention in CF with altered dietary or probiotic strategies. IMPORTANCE: While efforts have been focused on assessing the microbiome of pediatric and adult cystic fibrosis (CF) patients to understand how chronic colonization by these microbes contributes to pulmonary damage, little is known regarding the earliest development of respiratory and gut microflora in infants with CF. Our findings suggest that colonization of the respiratory tract by microbes is presaged by colonization of the gut and demonstrated a role of nutrition in development of the respiratory microflora. Thus, targeted dietary or probiotic strategies may be an effective means to change the course of the colonization of the CF lung and thereby improve patient outcomes.

Lactobacillus GG as an immune adjuvant for live-attenuated influenza vaccine in healthy adults: a randomized double-blind placebo-controlled trial.

BACKGROUND/OBJECTIVES: Live-attenuated influenza vaccine (LAIV) protects against influenza by mucosal activation of the immune system. Studies in animals and adults have demonstrated that probiotics improve the immune response to mucosally delivered vaccines. We hypothesized that Lactobacillus GG (LGG) would function as an immune adjuvant to increase rates of seroconversion after LAIV administration. SUBJECTS/METHODS: We conducted a randomized double-blind placebo-controlled pilot study to determine whether LGG improved rates of seroconversion after administration of LAIV. We studied 42 healthy adults during the 2007-2008 influenza season. All subjects received LAIV and then were randomized to LGG or placebo, twice daily for 28 days. Hemagglutinin inhibition titers were assessed at baseline, at day 28 and at day 56 to determine the rates of seroconversion. Subjects were assessed for adverse events throughout the study period. RESULTS: A total of 39 subjects completed the per-protocol analysis. Both LGG and LAIV were well tolerated. Protection rates against the vaccine H1N1 and B strains were suboptimal in subjects receiving LGG and placebo. For the H3N2 strain, 84% receiving LGG vs 55% receiving placebo had a protective titer 28 days after vaccination (odds of having a protective titer was 1.84 95% confidence interval 1.04-3.22, P=0.048). CONCLUSION: Lactobacillus GG is potential as an important adjuvant to improve influenza vaccine immunogenicity. Future studies of probiotics as immune adjuvants might need to specifically consider examining vaccine-naïve or sero-negative subjects, target mucosal immune responses or focus on groups known to have poor response to influenza vaccines.

Not just an individual journey: social aspects of recovery.

BACKGROUND: Recent literature on recovery describes the process as deeply personal and unique to each individual. While there are aspects of recovery that are unique to each individual, this article argues that focusing solely on these overlooks the fact that recovery unfolds within a social and interpersonal context. MATERIALS: Drawing from qualitative data, this article describes aspects of recovery that involve the contributions of others, the social environment and society. DISCUSSION: These aspects of recovery include relationships, adequate material conditions and responsive services and supports. CONCLUSION: The authors consider the implications of these social factors for transforming psychiatric research and theory as well as for recovery-orientated practice.

An investigation of adequate volume for the diagnosis of malignancy in pleural fluids.

OBJECTIVE: The cytological examination of pleural effusions is an important investigation in the diagnosis of malignancy. Maximizing the chances of identifying malignant effusions is therefore desirable. Recent Royal College of Pathologists guidelines, based on the British Society for Clinical Cytology codes of practice, have suggested that a minimum of 20 ml of pleural fluid is required for diagnostic purposes. METHODS & RESULTS: We examined 2155 pleural fluids received over a 6-year period in order to define a minimum required volume for adequacy. By examining the plateau phase of a graph of threshold volumes for initial samples received for each patient (n =1584) we determine that a minimum fluid volume of 25 ml is required and that more than 50 ml does not improve sensitivity. CONCLUSION: Between 25 and 50 ml of fluid are required for the adequate assessment of pleural effusions for malignancy.